Novel Protease-Activatable Chemerin-Derived Tracers for Molecular Imaging of Inflammation ABSTRACT Molecular imaging of inflammation allows for early identification of the pathogenic processes in a wide range of diseases and may lead to improved risk stratification of patients and monitoring the progression of disease or response to therapy. Thus, it represents a major driver of Precision Medicine, particularly with the growing development and applications of novel immunomodulatory therapeutics. However, limitations of the current molecular imaging tracers (e.g., low specificity and suboptimal kinetics) have been major challenges to successful targeted imaging of specific inflammatory processes. We have synthesized a prototype 99mTc-labeled high- affinity peptide, derived from the carboxy-terminus of a relatively recently identified chemokine, i.e., chemerin, which is internalized by macrophages expressing chemerin receptor 1 (also known as chemokine-like receptor 1 (CMKLR1). The goal of this proposal is to further advance this approach by the synthesis and optimization of novel activatable monomeric and cleavable multimeric tracers for in vivo imaging of chemerin-CMKLR1 axis in inflammation. Our central hypothesis is that the unique design of the activatable chemerin-derived imaging probes allows for accurate in vivo monitoring of inflammation, through taking advantage of: A) protease- mediated activatable mechanism, B) relatively restricted expression of CMKLR1 by immune cells, C) high affinity and subsequent receptor-mediated internalization of the C-terminal peptides, and D) enhanced kinetics of the multimeric peptides. We propose two Specific Aims: SPECIFIC AIM 1: To develop and optimize protease-activatable monomeric and cleavable multimeric chemerin- derived radiotracers. SPECIFIC AIM 2: To validate the accuracy of in vivo imaging using selected optimized tracers in murine models of sterile inflammation. We predict a high degree of species- independency of our tracers, which are derived from the highly conserved carboxy-terminus of chemerin. Thus, this approach may be applied in a variety of animal models and ultimately in humans. Our ultimate goal is to develop an innovative approach for targeted imaging of an important immunoregulatory pathway, i.e., chemerin-CMKLR1 axis, which may provide a venue for precision medicine in a variety of inflammatory diseases.